Page 1 of 11
Bioengineering 115: Cell Biology for Engineers
Laboratory 1: Cell Culture
Last updated: January 31, 2011
Lab 1: Cell Culture
The use of cultured cells grown
has several advantages over the use of an
system or an isolated tissue preparation in an experimental approach. This technique
allows the researcher to easily manipulate specific aspects of the cellular environment
and prevents exposure of an organism to toxic and other pharmacological agents.
Furthermore, the experimenter can selectively alter and study one variable at a time
without inducing secondary or tertiary effects of another system. Adding a chemical or
growth factor to a dish or flask of cells to determine its effect on those cells is
experimentally superior (in
ways) to simply injecting that factor or chemical into the
body. In the body, it is difficult to separate the direct effects of the factor or chemical on
the cells to the indirect effects.
Say you add factor X to cell type A in a dish, and observe the effects. At the same time,
you inject factor X into a lab animal, sacrifice the animal, and observe type A cells. With
the injection, all of the following (and more) are possible.
Factor X acts directly on cell type A.
Factor X effects cell type B, which releases factor Y, which
effects cell type
Factor X does nothing, because factor X is taken up by the liver before it can act
on cell type A.
Factor X does nothing, because it genuinely has no effect on cell type A.
When you use an isolated cell line, you remove a number of variables that make your
results more difficult to interpret. At the same time, your experiment becomes specific to
a particular cell, excluding practical contributions from the cell’s natural environment (the
Most cell types are grown as a single thickness cell layer (monolayer) or sheet attached
to glass or tissue culture treated plastic substrates. In order to keep a culture actively
growing, it is necessary to subdivide (e.g., split or passage) them at regular intervals.
Passaging initially involves the breakage of intercellular and cell-to-substrate connections
by the use or proteolytic enzymes (trypsin, collagenase, etc.). After the cells have been
disassociated into a suspension consisting of single cells, they are counted, diluted, and
transferred into fresh culture vessels. They re-attach, begin to grow and divide and after a
period of incubation (depending on the initial cell number, growth conditions, and the cell
line) they reach confluency. A passage of cells is considered confluent when the cells
cover the entire surface, and there is no room left for new cells. At this point the cells
must be passaged and subcultured (a portion of the cells used to create a new culture)
When dealing with cultured cells, it’s important to keep an eye on the growth media’s